#3230 characteristics of protein loss with medium cut-off membranes compared to high flux membranes

Abstract Background and Aims Medium cut-off membranes (MCO) have been introduced with the claim that larger pores enhance the removal of middle-sized uremic toxins. To date, no adequately powered randomized controlled trial has demonstrated that this membrane characteristic translates into improved mortality. A well-described consequence of larger pore sizes is a higher dialytic albumin loss [1]. In addition, other proteins could also be lost with MCO, including ones that are beneficial or even essential, e.g., to patients’ nutritional status. To better understand the spectrum of proteins cleared with MCO, we set up an ex vivo HD simulation system and compared protein loss characteristics between MCO and high-flux (HF) dialyzers. Method Over the course of an hour, a single reservoir of 2.5 liters EDTA human plasma was ultrafiltered using two Fresenius 2008T machines. One machine served the MCO dialyzer (Theranova 400), the other the HF dialyzer (Fresenius Optiflux F180NR). Ultrafiltrate (UF) was returned to the plasma reservoir (Figure 1A). Blood flow was 400ml/min, dialysate flow was zero, ultrafiltration rate was 13ml/min. UF samples were taken at ∼10 min intervals. Upon experiment completion, residual filtrate was collected from the dialysate compartment. Samples were stored at −80°C until analysis with Bradford protein assay, silver-stained SDS-Page, and Olink Proteomics (Watertown, MA). Selected samples underwent 3K MWCO protein concentration. Results Figure 1B shows the UF protein concentration at several time points. Total protein loss (in mg) during the one-hour simulation was 18 (HF) and 239 (MCO) in the UF, and 10 and 359 in the residual filtrate, respectively. SDS-PAGE protein gel revealed that HF filtrate contained less amount of protein than MCO filtrate when equal filtrate volumes were loaded (Figure 1C, lane 3 vs 2, lane 6 vs 5). Smaller proteins were more abundant in HF vs MCO filtrate when equal amounts of protein were loaded (lane 4 vs 2, lane 7 vs 5). UF from both membranes contained less proteins >100 kD than plasma (lane 1). Finally, UF (collected at 20 and 60 min) and residual filtrate were concentrated to ∼1mg/ml and analyzed using targeted biomarker analysis with Olink® Target 96 Organ Damage panel. Ten of the 92 panel proteins were above the limit of detection (LOD; Table 1). The MCO-to-HF protein concentration ratio in equal filtrate volumes ranged from 1.7 to 13.1. Conclusion Our results show a higher amount of protein loss with MCO compared to HF. The HF filtrate protein distribution is skewed towards lower molecular weights compared to MCO filtrate. All 10 proteins identified were more abundant in MCO filtrate compared to HF filtrate. Further identification and quantification of protein loss during hemodialysis will improve our understanding of the clinical impact of different dialysis membranes.